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==PCR amplification of cosmic probe== PCR reaction system x48 H2O 50ul 2400ul SYBR green (50x) 0.4ul 19.2ul primer mix (100uM) 0.2ul 9.6ul Taq master mix(2x) 50ul 2400ul probe template (cosmic) 0.05ul 2.4ul 94c 2min -> 94c 30sec -> 60C 2min -> 72C 1min-> Plate read -> (94c 30sec -> 60C 2min -> 72C 1min)x 20 cycles -> 72C 5min -> 15C hold Purify the product using ethanol purification: 1. pool the 24 reactions into one 16 mL tube, add 240 uL 3M NaOAC, 8 uL GlycoBlue, 6 mL 100% Ethanol. 2. Store at -70C for more than 20 min. 3. Spin at 4000rpm for 15 min at 4C. 4. Remove all liqid, you should see a blue pallette at the bottom of the tube. 5. Add 750 uL cold 75% Ethanol, piette well and transfer to a 1.6 mL tube. 6. Spin at 10000 rpm for 5 min at 4C. 7. Remove all liquid, let the pallette dry for 5 min, resuspend the DNA with 100 uL dH2O.
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